Folate metabolism is required for the synthesis of nucleotides (purines and dTMP) and S- adenosylmethionine (AdoMet). Disruption of folate metabolism affects AdoMet and dTMP syntheses and thereby influences AdoMet-dependent methylation reactions and uracil content in DNA. Both DNA uracil and methylation affect DNA stability, and chromatin methylation regulates the expression of many genes. It is not known if the associations between disruptions in folate metabolism and pathologies (including certain cancers, cardiovascular disease) and developmental anomalies (including neural tube defects (NTDs)) result from altered AdoMet synthesis and/or dTMP synthesis. Recently, we demonstrated that the enzyme cytoplasmic serine hydroxymethyltransferase (cSHMT) is a metabolic switch that regulates the flux of folate-activated one-carbon units between the dTMP and AdoMet biosynthetic pathways. Reduced expression of cSHMT, as observed in both cSHMT and cSHMT-/- mice, induces embryonic NTDs, sensitizes the animals to colon cancer, affects uracil content in DNA and increases the AdoMet/AdoHcy ratio. This is the only mouse model to exhibit NTDs as a result of the disruption of a folate-dependent enzyme, and thereby enables elucidation of the mechanisms underlying folate-responsive NTDs. CSHMT is expressed in tissues known to be associated with folate-related pathologies/developmental anomalies including NTDs. The expression and activity of cSHMT is dynamically regulated by several nutrients and therefore the cSHMT has the potential to contribute to the etiology of folate-related pathologies and may be a target for prevention through diet. The contributions of diet, embryonic and maternal cSHMT genotype to NTD occurrence will be determined, and the metabolic role of cSHMT in NTD etiology will be elucidated. The specific aims are: 1) to determine the gene-nutrient interactions that increase risk for NTDs in mice deficient in cSHMT. 2) to determine the metabolic defect associated with neural tube closure defects. 3) to elucidate the contribution of cSHMT SUMOylation to NTD frequency. 4) to determine if cSHMT and MTHFR interact to increase risk for folate-responsive NTDs. The principle hypotheses to be tested are that: 1) cSHMT expression contributes to folate-responsive NTD risk by altering folate metabolism. 2) impairments in cSHMT SUMOylation affect dTMP and AdoMet synthesis and risk for NTDs. 3) the cSHMT and MTHFR genes interact to increase risk for folate-responsive NTDs. PUBLIC HEALTH RELEVANCE: The long-term goals of this project is to determine the mechanisms underlying neural tube closure defects and the role of cSHMT in NTD prevention. Folate-related pathologies, including certain cancers, and developmental defects, including neural tube defects, are common and complex disorders involving gene nutrient interactions but the underlying mechanisms are not established. Folate-fortification of the US food supply was implemented to reduce incidence of birth defects, yet concerns remain regarding the effects of folate fortification on cancer incidence. The studies outlined in this proposal will investigate the mechanisms of folate-related birth defects and role of folate and other dietary components in preventing these defects in the first mouse model of folate-related birth defects resulting from disrupted folate metabolism.